This renewal application proposes extensive and innovative studies focusing on the molecular genetics of the cutaneous basement membrane zone (BMZ) towards delineating the molecular basis of various forms of epidermolysis bullosa (EB) and other selected genodermatoses affecting the epidermis. The proposed studies are designed to test the hypothesis that genetic lesions in structural genes expressed in the epidermis underlie variants of these diseases, and that the precise phenotype and mode of inheritance depend on the types and combinations of specific mutations in distinct genes. This application is based on solid progress in this project, including (a) expansion of the molecular basis of the recessive dystrophic forms of EB allowing refinement of genotype/phenotype correlations; (b) identification of novel and de novo COL7A1 mutations in dominant DEB, with an impact on genetic counseling of the families at risk of recurrence; (c) identification of a large number of novel and recurrent mutations both Herlitz and non-Herlitz junctional EB; (d) identification of uniparental disomy of chromosome 1 as a novel mechanism for H-JEB; (e) demonstration in mutations in the genes ITGA6 and ITGB4 encoding alpha-6-beta-4 integrins subunit polypeptides in EB with pyloric atresia,; (f) cloning of the human plectin gene and demonstration of mutations in EB with late-onset muscular dystrophy; (g) cloning of mouse type VII collagen and desmoglein 3 genes with development of "knock-out" mice with blistering phenotype; (h) identification and characterization of several novel genes expressed into the epidermis, including periplakin, ladinin, and desmo-15; (i) refinement of RNA- DNA chimeric oligonucleotide technology for repair of the mutated genes in heritable skin diseases. This proposal details continuation of concentrated, multi-disciplinary studies in five highly interdependent projects: Project 1, "Molecular Genetics of EB and Other Heritable Disorders of the Cutaneous BMZ and Epidermis," will provide precise information on the specific mutations in the gene/protein system that are at fault in various forms of EB and other epidermal heritable disorders. Project 2, "Identification and Characterization of Candidate Genes/Protein Systems Expressed in the Skin," will provide new gene probes and information about novel genes as potential candidate genes for epidermal genodermatoses. Project 3, "Consequences of the Mutations at the Protein Structure/Function Level" will examine the structural and functional alterations that result from distinct mutations in the candidate genes, utilizing computer modeling and monitoring functional interactions in biosensor analysis system. Project 4, "Development and Testing of Animal Models for EB," will generate novel animal models for EB. Project 5, "Development of Non-Viral Gene Therapy for Cutaneous Diseases," will concentrate on testing gene therapy approaches utilizing RNA/DNA chimeric oligonucleotide strategies. These multidisciplinary studies are expected to provide precise information of critical importance for translational applications towards development of refined classification, genotype/phenotype correlations, basis for genetic counseling, and prenatal testing, as well as providing the basis for novel gene therapy approaches for this devastating group of skin diseases.